Study On Optimalize Of Maize Transformation System And Transformation Of PwHAP5

Maize (Zea.mays) is the third-largest food crop on the world. It's an important food crops and industry materials. Because the planting area of maize is wide and large in the world, and the maize is the high output and good adaptability, it is important to research genetic transformation of maize and improve corn varieties. Therefore, it received a wide attention on the world.1. The maize self-bred line H99 is the material of this study. The callus is induced from young embryo which is self pollinated. This study find that the age of embryo is significant for callus.It is prejudice for callus induction organization, if the young embryo is too small or too big. It is easy to induce callus organization when the embryos is 1.3 ~ 1.7mm, and the induction rate is the highest. The concentration of 2,4-D has significant influence on callus induction during the process of young embryo induction.The better concentration of 2,4-D for maize callus induction organization is 3mg /L.2. The state of transgenic receptor system directly affects the efficiency of conversion rate and transformation. There is three types of maize callus, and theâ…¡type of embryogenic callus tissue is the most favourable for transformation. This study got the formula of culture medium of transformation from other types of callus toâ…¡type callus. Theâ…¡type of embryogenic callus induction medium formula is 2MS basic salt, 600mg/L L-proline, 200mg/L L-asparagine, 400mg/L CH, 3.3 mg/L Dicamba, 10.0 mg/L AgNO3 and 30g/L sucrose.3. Extract pollen's mRNA of Picea wilsonii after incubated 12h. the ORF sequence of HAP5 was got. It is transcription factor of HAP class by reverse transcription, named PwHAP5. The sequence of ORF of the transcription factors PwHAP5 has 603 nucleotide, coding 201 amino acids. The important structures of transcription factor PwHAP5 of Picea wilsonii are more conservative, which is like arabidopsis, rice, yeast and human. The genetic relationship between PwHAP5 and NF-YC2 of Arabidopsis is closed. The expression protein of this gene is without transmembrane structure.4. PwHAP5 has transformed to tobacco and corn by Agrobacterium-mediated technique. Got two transgenic tobacco and three transgenic maize, and extracte the DNA of transgenic plants. Use PwHAP5 specificity primer to amplificate, objective strip was got.